An optical arrangement of the kind recited above is known, for example, from DE 10 2007 047 464 A1. The known optical arrangement comprises an illumination device for generating a light sheet illuminating a sample; and a detection device, comprising a detector, for detected light proceeding from the sample. In order to generate the light sheet necessary for SPIM, the known SPIM arrangement comprises suitable lenses in the illumination beam path. Also known, from DE 20 2011 110 077 U1, is a SPIM arrangement in which a light sheet is generated via a mirror arrangement at the detection objective.
One known area of utilization of SPIM technology is the field of fluorescence microscopy, fluorophores in the sample being excited with laser light. With SPIM technology, excitation takes place only in the plane of the light sheet. Damage to the sample from illumination light in other planes is thereby avoided.
In a SPIM arrangement, a light sheet is generated either by using an extended focus with the aid of a scanning device in order to illuminate a plane, or by using a cylindrical lens. Because it is usually desirable to illuminate a large area of objects, or large objects, the illumination aperture (NA) must be a low as possible. A consequence of this, however, is that the light sheet is very thick.
In the case of a long light sheet—approx. 300 μm for illumination with a 2.5× objective having an NA of 0.07—the light sheet thickness is usually greater than the depth of focus of the detection objective. Image data from the other two planes of the light sheet located above and below the actual depth of focus (unsharp regions above and below the actual depth of focus) cannot be used. This is very ineffective, and has the additional disadvantage that the sample regions located above and below the actual depth of focus are illuminated entirely needlessly, and as a result can possibly be damaged.